Methanediphosphonic acid derivative, process for production thereof and use for pharmaceuticals

ABSTRACT

A methane diphosphonic acid derivative represented by the general formula (I): ##STR1## wherein, X and Y are defined in the specification; m represents an integer of 0 to 3; n represents an integer of 0 to 4; and each X of the (X) m  and each Y of the (Y) n  may be either identical or different; . . . represents a double bond or single bond; A is --(CH 2 )a--(D)b--(CH 2 )c-- (wherein D is sulfur, oxygen, NH, alkyl-substituted N, or CH 2 , a and c are integers of 0 to 10 and b is 0 or 1), or --(CH═CH)d═CH= (wherein d is an integer of 0 to 2, and B does not exist when A represents --CH═CH)d--CH═), B refers to a hydrogen atom, alkyl group, amino group, monoalkylamino group, dialkylamino group, acylamino group, hydroxyl group, alkoxy group, trialkylsiloxy group or acyloxy group, and each of R 1 , R 2 , R 3  and R 4  is hydrogen atom, straight or branched alkyl group having 1 to 7 carbon atoms, or pharmacologically allowed cation, and these may be identical or different, as described. The compounds of the present invention have excellent IL-1 inhibitory action, antioxidative action and bone resorption inhibitory action, and are useful as an antiinflammatory drug, antirheumatic drug, or autoimmune disease drug.

TECHNICAL FIELD

The present invention relates to a novel methanediphosphonic acidderivative which inhibits actions of interleukin-1 that mediatespyrexia-inducing reactions, inflammation-inducing reactions, activatesvarious blood cells and has bone destructive action, whilesimultaneously having an action to inhibit active oxygen that cause celldamage and fat denaturation, as well as an action to inhibit bonedestruction during osteoporosis and chronic articular rheumatism.

BACKGROUND ART

Many of the diphosphonic acid compounds that have been developed mainlyas drugs for treatment of bone metabolic diseases thus far have anaction to inhibit bone destruction, and have been expected to inhibitbone destruction during the occurrence of arthritis such as chronicarticular rheumatism. Although compounds having a diphosphonic acidstructure are disclosed in Japanese Unexamined Patent Publication(Kokai) No. 59-42395, Japanese Unexamined Patent Publication (Kokai) No.2-22285, Japanese Unexamined Patent Publication (Kokai) No. 3-77894 andJapanese Unexamined Patent Publication (Kokai) No. 60-174792, thesediphosphonic acid compounds are primarily focused on inhibition of boneresorption. Although these compounds are effective as therapeutic drugsfor bone metabolic disorders, they are still not adequate for treatmentof chronic articular rheumatism.

In order for diphosphonic acid compounds to be used in the treatment ofchronic articular rheumatism and so forth, a new drug is desired that,in addition to having an action to inhibit bone resorption, also hasother, more effective actions, including inhibition of Interleukin-1(abbreviated as IL-1), which is a mediator of inflammations, as well asinhibition of cell damage caused by activated neutrophils andmacrophages.

IL-1 is known to be a mediator involved in pyrexia and inflammation, andits inhibitory agent is expected to be useful as an antiinflammatorydrug. However, similar to many other cytokines, IL-1 is considered tomainly act locally. Although numerous substances have been reported toinhibit IL-1 in vitro, antiinflammatory drugs having action that allowsadequate improvement of the disease state by actually inhibiting IL-1 invivo have not yet been developed. In addition, invasion of activatedneutrophils and macrophages at the site of inflammation have beenobserved during inflammations. Although the active oxygen produced bythese blood cells has an action of heterogenous digestion, in caseswhere an inflammation has become chronic, these cells are known todamage normal tissue as well. Thus, compounds having both an IL-1inhibiting action and antioxidative action are considered to be usefulas not only antiinflammatory drugs, but also against autoimmune diseasessuch as chronic articular rheumatism, as well as organ disorders, suchas those in the brain and liver, which occur during ischemia.

DISCLOSURE OF THE INVENTION

The present inventors conducted research on diphosphonic acid compoundsthat demonstrate excellent anti-inflammatory action, by giving to adiphosphonic acid derivatives not only action as therapeutic drugs fortreatment of bone metabolic diseases, but also IL-1 inhibitory actionand antioxidative action. During the course of this research, it wasfound that, if a naphthalene skeleton is given to a phosphonic acidstructure, IL-1 inhibitory action and antioxidative action are providedthat are not found in existing drugs.

The object of the present invention is to provide a useful, novelcompound having an action to inhibit Interleukin-1, antioxidativeaction, and an action to inhibit bone destruction.

Namely, the present invention relates to the methane diphosphonic acidderivatives shown in general formula (I): ##STR2## [wherein, X and Yrepresent substituent groups on the naphthyl group, and represent ahalogen atom, nitro group, nitrile group, alkyl group, alkoxy group,trifluoromethyl group, the group: ##STR3## (wherein Z¹ and Z² representindependently hydrogen atom or alkyl group, or Z¹ and Z² together mayform a ring composed of carbon atoms or a ring composed of carbon atomsand hetero atom), the group: ##STR4## (wherein Z¹ and Z² are the same asabove, and Z³ represents oxygen or sulfur), thiol group, hydroxyl group,alkylthio group, arylthio group, acyloxy group, acylamino group,acylthio group, acyl group, alkenyl group, aryl group, cycloalkyl group,COOH group or COO-alkyl group; m represents an integer of 0 to 3, nrepresents an integer of 0 to 4, and each X of the (X)_(m) and each Y ofthe (Y)_(n) may be either identical or different; . . . represents adouble bond or single bond; A is --(CH₂)a--(D)b--(CH₂)c-- (wherein D issulfur, oxygen, NH, alkyl-substituted N, or CH₂, a and c are integers of0 to 10 and b is 0 or 1), or --(CH=CH)d--CH= (wherein d is an integer of0 to 2, and B does not exist when A represents --CH═CH)d--CH═); Brepresents a hydrogen atom, alkyl group, amino group, monoalkylaminogroup, dialkylamino group, acylamino group, hydroxyl group, alkoxygroup, trialkylsiloxy group or acyloxy group; and R¹, R², R³ and R⁴ arehydrogen atom, straight or branched chain alkyl group having 1 to 7carbon atoms, or pharmacologically acceptable cation, and may beidentical or different], a process for production of said derivative, aswell as its pharmaceutical applications, such as antiinflammatory drugs,antirheumatic drugs, bone metabolic disease drugs, an Interleukin-1inhibitor, an antioxidant and a bone destruction inhibitor, each havingsaid derivative as its active ingredient.

DETAILED DESCRIPTION

In the case of an unsubstituted naphthyl group, a 1-naphthyl group or2-naphthyl group is represented. In the case of a naphthyl group having1 or more substituent groups, a 1-naphthyl group having substituentgroup(s) at the 2 to 8 positions, or a 2-naphthyl group havingsubstituent group(s) at the 1 position or 3 to 8 positions, isrepresented. In the case the naphthyl group has substituent group(s),preferable location(s) of the substituent group(s) are the 2 and/or 4positions when substituted with X and the 5 and/or 6 and/or 8 positionswhen substituted with Y, in the case of a 1-naphthyl group; and the 1and/or 4 positions when substituted with X and the 5 and/or 6 and/or 8positions when substituted with Y, in the case of the 2-naphthyl group.Those halogen atoms used as substituent groups X and Y are fluorine,chlorine, bromine and iodine. Alkyl group (alkyl group indicatedhereinafter also has the same meaning) is straight or branched chainalkyl group having 1 to 7 carbon atoms, and for example, methyl, ethyl,propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, hexyl, cyclohexylgroup etc. Alkoxy group is those having 1 to 7 carbon atoms, forexample, methoxy, ethoxy, propoxy, isopropoxy, butoxy group etc.Examples of the group: ##STR5## (wherein the alkyl groups of Z¹ and Z²are the same as described above) include amino, methylamino, ethylamino,propylamino, butylamino, dimethylamino, diethylamino, pyrrolidino,piperidino, morpholino, thiomorpholino group etc. Examples of the group:##STR6## (wherein Z¹, Z², and Z³ are the same as described above)include carbamoyl, thiocarbamoyl, N-methylaminocarbonyl,N,N-dimethylaminocarbonyl, piperidinocarbonyl, pyrrolidinocarbonyl,morpholinocarbonyl group etc. Examples of alkylthio group (wherein thealkyl moiety is the same as the alkyl group described above) includemethylthio, ethylthio, propylthio, isopropylthio, cyclopentylthio,cyclohexylthio group etc. Arylthio group is preferably those having 6 to15 carbon atoms, examples of which include phenylthio, substitutedphenylthio groups etc. The acyl (group) of acyloxy, acylamino, acylthioand acyl groups is straight or branched chain group having 2 to 7 carbonatoms, examples of which include acetyl, propanoyl, butanoyl groups etc.Alkenyl group is straight or branched alkenyl group having 2 to 7 carbonatoms, examples of which include vinyl, allyl, 2-propenyl, isopropenyl,butenyl, pentenyl groups etc. Aryl group is preferably those having 6 to15 carbon atoms, examples of which include phenyl, substituted phenyl,naphthyl groups etc. Cycloalkyl group is those having 3 to 8 carbonatoms, examples of which include cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl groups etc. Examples of COO alkyl group (wherein the alkylmoiety is the same as the alkyl group previously described) includemethoxycarbonyl, ethoxycarbonyl, propoxycarbonyl groups etc.

In the case where A is --(CH₂)a--(D)b--(CH₂)c--, and . . . represents asingle bond, D is sulfur, oxygen, NH, alkyl-substituted N (wherein alkylis a straight or branched chain alkyl group having 1 to 7 carbon atoms)or CH₂ ; a and c are integers of 0 to 10, and b is 0 or 1 (provided thata=c=0 when b=0). However, more preferably, a, b and c are independently0 or 1.

Alternatively, in the case where A is --(CH=CH)d--CH=, . . . is a doublebond, d is an integer of 0 to 2 and B does not exist.

Moreover, in the case where B is other than hydrogen atom and an alkylgroup, and D is CH₂ and b is an integer other than 1, those compoundswherein c=0 are not preferable since they are chemically unstable.However, even in this case, those compounds wherein a=b=c=0 arepreferable because they are stable.

Particularly preferable specific examples include those in which A is S,O, NH, CH═, CH₂, CH₂ S, CH₂ O, CH₂ NH, CH₂ CH₂, SCH₂, SCH₂ CH₂, SCH₂ CH₂CH₂, OCH₂ and NHCH₂. In addition, those compounds wherein the naphthylgroup is directly bonded to the carbon atom of the methane diphosphonicacid without through the A (namely, the case wherein a=b=c=0) are alsoincluded.

Alkyl moiety in the cases wherein B is an alkyl group, monoalkylaminogroup, dialkylamino group, alkoxy group and trialkylsiloxy group is thesame as the alkyl group described above, and the acyl moiety ofacylamino and acyloxy groups is the same as the acyl groups describedabove.

Typical examples of the alkyl group of R¹, R², R³ and R⁴ include methyl,ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl groups andthe like.

In the case where R¹, R², R³ and R⁴ are hydrogen atoms, the phosphonicacid moiety of formula (I) can form a salt with inorganic or organicbase. Pharmacologically allowed cations in this case refer to metalcations and ammonium ions NR₄ (wherein R is the same as the alkyl groupand hydrogen atom of R¹ to R⁴). Particularly preferable metal cationsinclude cations of alkaline metals, such as lithium, sodium, potassiumetc, as well as cations of alkaline earth metals, such as magnesium,calcium etc. However, the cation of other metals, such as aluminum,zinc, iron etc, are also included in the present invention. Examples ofammonium ions include ammonium ions of ammonia, primary amines,secondary amines and tertiary amines, as well as quaternary ammoniumions. Examples thereof include ammonium ions of ammonia, methylamine,dimethylamine, trimethylamine, ethylamine, diethylamine, triethylamine,propylamine, dipropylamine, isopropylamine, diisopropylamine,butylamine, dibutylamine, isobutylamine, t-butylamine, monoethanolamine,diethanolamine, triethanolamine etc, as well as tetramethylammonium,tetraethylammonium ions etc. Cations of sodium, potassium, ammonia andalkylamines are particularly preferable. In addition, in R¹ to R⁴, thecations may be identical or different, and those containing a mixture ofhydrogen atom and cation are also included in the present invention,examples of which include monocationic salts, dicationic salts andtricationic salts. The methane diphosphonic acid derivatives shown inthe general formula (I) is preferably those wherein R¹ to R⁴ are allhydrogen atoms, those wherein three of R¹ to R⁴ are hydrogen atoms,while the other is sodium, those wherein three of R¹ to R⁴ are hydrogenatoms while the other is ammonium, those wherein two of R¹ to R⁴ arehydrogen atoms and the remaining two are sodium, or those wherein two ofR¹ to R⁴ are hydrogen atoms while the remaining two are ammonium.

The methane diphosphonic acid derivatives of the present invention canbe produced by a process resembling a known process in said field. Forexample, one of the methane diphosphonic acid derivatives of formula (I)of the present invention (in the case where B is a hydrogen atom) can beproduced by the process indicated by the following reaction formula:##STR7##

The starting substance that is used is a lower alkyl ester of methanediphosphonic acid (II) (wherein the lower alkyl is a straight orbranched chain alkyl having 1 to 7 carbon atoms). The correspondingmetallized methane diphosphonic ester (y) is formed by reacting theabove starting substance with a base such as sodium hydride or alkyllithium. This is then reacted with various naphthyl-A group introducingagents (wherein naphthyl is: ##STR8## (wherein X, Y, m and n are thesame as previously described), and A is the same as previously defined)to form compound (VI). Examples of naphthyl-A group introducing agentsthat are used include halogen compounds such asnaphthyl--(CH₂)a--(D)b--(CH₂)c-halogen.sub.,[naphthyl--(CH₂)a-S-halogen, etc., or a disulfide ofnaphthyl--(CH₂)a-S]₂ (wherein D, a, b and c are the same as describedabove).

The reaction temperature and reaction time vary according to thereagents used. For example, the reaction temperature is between -78° C.and the boiling point of a solvent or solvent mixture, and the reactiontime is from 10 minutes to several days.

An example of another synthetic process for a derivative of the methanediphosphonic acid of the general formula (I) is shown by the followingreaction formula: ##STR9## and the lower alkyl ester (II) of methanediphosphonic acid are reacted in as a condensation reaction in thepresence of titanium tetrachloride and a tertiary amine such asN-methylmorpholine to obtain compound (VII). Moreover, the double bondthat is formed is reduced to obtain compound (VIII).

A methane diphosphonic acid derivative, wherein R¹ through R⁴ arehydrogen atoms, can be obtained from a methane diphosphonic acidderivative wherein R¹ through R⁴ are alkyl groups (phosphonic ester) byhydrolysis and so forth. For example, a phosphonic ester is hydrolyzedeither by reacting with an acid such as hydrochloric acid, or treatingwith trimethylsilylbromide followed by water or alcohol. The methanediphosphonic acid thus obtained can be converted to one of salts thereofby a known process.

In addition, compounds wherein one to three of R¹ through R⁴ are alkylgroups (partial esters of methane diphosphonic acid) obtained by partialhydrolysis of a methane diphosphonic ester or partial esterification ofmethane diphosphonic acid are also included in the present invention.

In addition, although the P═O bonds in the majority of the methanediphosphonic acid derivatives of the present invention exist in the ketoform, there are cases wherein a portion of these bonds may exist in theenol form depending on the chemical properties of the compound itself,solvents and temperature. However, these compounds are also included inthe compounds of the present invention.

In addition, in all the reactions, in the case where reactivesubstituent groups and reactive functional groups for reactions otherthan the desired reaction are contained, these substituent groups andfunctional groups must be protected in advance by reagents that allowthem to be easily removed.

Those diseases at which compounds of the present invention are directedare inflammatory diseases, pain diseases, skin diseases, respiratoryorgan diseases, liver diseases, infections, autoimmune diseases,ischemic organ disorders and bone metabolic diseases. For example, thepresent invention provides a drug having excellent therapeutic andpreventive activity against (chronic) articular rheumatism, rheumatoidpolyarthritis, osteoarthritis, scapular periarthritis, neck-shoulder-armsyndrome, intervertebral disk disorders, lumbago, tendinitis andperitendinitis, arthrosteitis, stiff and painful shoulder, fibrositis,muscle pain, neuralgia, gout, post-surgical and posttraumaticinflammation and swelling (antiinflammatory drugs, antirheumatic drugs,antiarthritic drugs, analgesics and antipyretics), or psoriasis, asthma,pulmonary sarcoidosis, viral hepatitis, human immunodeficiency viralinfections, protozoan infections, ischemic heart disease, ischemicencephalopathy, ischemic hepatitis, arteriosclerosis, osteoporosis,Paget's disease, Bechterew's disease, hypercalcemia and ectopicossification (bone metabolic disease drugs).

In the case of using the novel methylene or methane diphosphonic acidderivatives of the present invention in the applications of the presentinvention listed above, said derivatives can be provided for use eitheras such is or in the form of pharmaceutical compositions mixed withknown pharmacologically acceptable carriers, vehicles and so forth.Above-mentioned derivatives may either be given by oral administrationin the form of tablets, capsules, powders, granules or pills, or byparenteral administration in the form of injections, syrups, ointmentsand suppositories. Although the dose varies according to the patient,administration route and symptoms, it is approximately 1 mg to 5 g, andpreferably 1 mg to 2 g. This dose may given either orally orparenterally once or several times per 1 day, or once per 1 to 7 days.

EXAMPLES

A more concrete explanation of the present invention will now beprovided with reference to the Examples.

Example 1 Tetraethyl 2-naphtbylthiomethanediphosphonate (1) ##STR10##

Under an argon atmosphere, a solution of 10.09 g (35 mmol) oftetraethylmethanediphosphonate in dry tetrahydrofuran (100 ml) wascooled to -78° C., and then 22.01 ml (35 mmol) of a solution of n-butyllithium in hexane [1.59 mol/l] was added thereto, and the mixture wasstirred for 30 minutes. Next, a solution of 11.15 g (35 mmol) of2,2'-dinaphthyl disulfide in dry tetrahydrofuran (75 ml) was added tothe mixture, which was then warmed to room temperature, and then stirredfor 16 hours. The resulting mixture was poured into ice water andneutralized with 6N hydrochloric acid, and then extracted with ethylacetate (3×150 ml). The organic layer was washed with water andsaturated saline and then dried over anhydrous magnesium sulfate. Next,the solvent was distilled off under reduced pressure, and the resultingresidue was purified by column chromatography (developing solvent -ethanol:ethyl acetate=5:95) to obtain 8.82 g of the title compound as acolorless oil. The yield was 57%.

¹ H-NMR (CDCl₃) [ppm]: 1.33 (t, J=7 Hz, 12H), 3.55 (t, J=21 Hz, 1H),4.00-4.55 (m, 8H), 7.46-7.52 (m, 3H), 7.76-7.83 (m, 3H), 8.07 (S, 1H)

IR (KBr) [cm⁻¹ ]: 2984, 1626, 1589, 1502, 1392, 1257, 1029, 975 MASS(FAB) m/z: 447 (M+H)⁺ EA (as C₁₉ H₂₃ O₆ P₂ S) Calculated (%): C 51.12 H6.33 Found (%): C 51.10 H 6.29

Example 2 2-naphthylthiomethane Diphosphonic Acid (2) ##STR11##

Under an argon atmosphere, to a solution of 8.04 g (18 mmol) oftetraethyl 2-naphthylthiomethane-diphosphonate obtained in Example 1 indry methylene chloride (100 ml) was added dropwise 27.56 g (180 mmol) oftrimethylsilyl bromide at room temperature and then the mixture wasstirred at room temperature for 72 hours. After the solvent and theexcess trimethylsilyl bromide were distilled off under reduced pressure,the resulting residue was dissolved in a mixed solvent ofwater:methanol=5:95, and the solution was heated to reflux for 30minutes, and the solvent was again distilled off under reduced pressure.The resulting residue was crystallized from acetone/methylene chlorideand the obtained crystals were recrystallized from the same solvent toobtain the title compound as white crystals. The yield was 86%.

m.p.: 218.5°-219.5° C. (dec)

¹ H-NMR (CD₃ OD ) [ppm]: 3.51 (t, J=21 Hz, 1H ), 7.42-7.51 (m, 2H),7.65-7.70 (m, 1H), 7.74-7.87 (m, 3H), 8.10-8.12 (m, 1H)

IR (KBr) [cm⁻ ]: 2926, 1657, 1638, 1620, 1151, 973, 812 MASS (FAB) m/z:335 (M+H)⁺ EA (as C₁₁ H₁₂ O₆ P₂ S) Calculated (%): C 39.53 H 3.63 Found(%): C 39.62 H 3.70

Example 3 Tetraethyl 6-methoxy-2-naphthylthiomethanediphosphonate (3)##STR12##

(a) 6,6'-Dimethoxy-2,2'-dinamhthyl disulfide

Under an argon atmosphere, a solution of 10.00 g (28.5 mmol) of6,6'-dihydroxy-2,2'-dinaphthyl disulfide in dry dimethylformamide (250ml) was cooled to -23° C., and then 3.42 g (85.5 mmol) of sodium hydride(60% dispersion in mineral oil) was slowly added thereto, and themixture was stirred until the generation of hydrogen ceased. 12.14 g(85.5 mmol) of methyl iodide was added to the mixture, which was thenallowed to stand at room temperature and stirred for 2 hours. Theresulting mixture was poured into ice water, and extracted with ethylacetate (3×150 ml). The organic layer was washed with water andsaturated saline and then dried over anhydrous magnesium sulfate. Thesolvent was distilled off under reduced pressure. The obtained crystalswere recrystallized from ethyl acetate/petroleum ether to obtain 9.92 gof 6,6'-dimethoxy-2,2'-dinaphthyl sulfide as orange crystals. The yieldwas 92%.

m.p.: 125°-126° C.

¹ H-NMR (CDCl₃) [ppm]: 3.89 (s, 6H), 7.00-7.25 (m, 4H), 7.45-7.75 (m,6H), 7.85-7.95 (m, 2H)

(b) Tetraethyl 6-methoxy-2-naphthylthiomethanediphosphonate

Following the same method as described in Example 1, from 10.09 g (35mmol) of tetraethyl methanediphosphonate and 13.25 g (35 mmol) of6,6'-dimethoxy-2,2'-dinaphthyl disulfide was obtained 11.34 g of thetitle compound as a pale yellow oil. The yield was 68%.

¹ H-NMR (CDCl₃) [ppm]: 1.33 (t, J=7 Hz, 6H), 1.35 (t, J=7 Hz, 6H), 3.48(t, J=22 Hz, 1H), 3.91 (s, 3H), 4.00-4.45 (m, 8H), 7.00-7.30 (m, 2H),7.50-7.80 (m, 3H), 8.00-8.10 (m, 1H)

IR (KBr) [cm⁻¹ ]: 2984, 2936, 2910, 1628, 1593, 1390, 1259, 1214, 1023,975 MASS (FAB) m/z: 477 (M+H)⁺ EA (as C₂₀ H₃₀ O₇ P₂ S) Calculated (%): C50.42 H 6.36 Found (%): C 50.65 H 6.42

Example 4 6-Methoxy-2-naphthylthiomethanediphosphonic Acid (4) ##STR13##

Following the same method described as in Example 2, 7.15 g (15 mmol) oftetraethyl 6-methoxy-2-naphthylthiomethanediphosphonate obtained inExample 3 in dry methylene chloride was treated with trimethylsilylbromide, and then hydrolyzed to obtain 4.21 g of the title compound aswhite crystals. The yield was 77%.

m.p.: 234°-235° C. (dec)

¹ H-NMR (CD₃ OD) [ppm]: 3.37 (t, J=21 Hz, 1H), 3.90 (s, 3H), 7.12-7.23(m, 2H), 7.65-7.75 (m, 3H), 8.05-8.09 (m, 1H)

IR (KBr) [cm⁻¹ ]: 2920, 1628, 1466, 1214, 1125, 994, 924, 911 MASS (FAB)m/z: 365 (M+H)⁺ EA (as C₁₂ H₁₄ O₇ P₂ S) Calculated (%): C 39.57 H 3.88Found (%): C 39.55 H 3.79

Example 5 Tetraethyl 6-hydroxy-2-naphthylthiomethanediphosphonate (5)##STR14##

(a) 6,6'-Di(t-butyldimethylsiloxy)-2,2'-dinaphthyl disulfide

Under an argon atmosphere, a solution of 10.00 g (28.5 mmol) of6,6'-dihydroxy-2,2'-dinaphthyl disulfide and 9.70 g (140 mmol) ofimidazole in dimethylformamide (150 ml) was cooled to 0° C., and asolution of 12.89 g (85.5 mmol) of t-butyldimethylchlorosilane in drydimethylfomamide (50 ml) was added thereto, and the mixture was warmedto room temperature and then stirred for 3 hours. The resulting mixturewas poured into ice water, and extracted with ethyfl acetate (3×150 ml).The organic layer was washed with water and saturated saline and thendried over anhydrous magnesium sulfate. The solvent was distilled offunder reduced pressure. The obtained residue was purified by columnchromatography (developing solvent - ethyl acetate:n-hexane =5:95) toobtain the title disulfide compound as a pale yellow oil. The yield was99%.

¹ H-NMR (CDCls) [ppm]: 0.23 (s, 6H), 1.00 (s, 9H), 6.95-7.20 (m, 2H),7.45-7.70 (m, BH), 7.85-7.95 (m, 1H)

(b) Tetraethyl 6-hydroxy-2-naphthylthiomethanediphosphonate

Following the same method as described in Example 1, a reaction between10.09 g (35 mmol) of tetraethyl methanediphosphonate and 13.25 g (35mmol) of 6,6'-di(t-butyldimethylsiloxy)-2,2'-dinaphthyl disulfide wascarried out. After evaporation of the solvent of the reaction mixture,the resulting residue was dissolved in a mixed solvent of 6Nhydrochloric acid:methanol=1:20, heated at 50° C. for 30 minutes, andthe solvent was again distilled off under reduced pressure. The obtainedresidue was purified by column chromatography (developing solvent -ethanol:ethyl acetate=5:95) to obtain 10.02 g of the title compound as apale yellow oil which slowly crystallized. The yield was 62%.

m.p.: 85.5°-86.5° C.

¹ H-NMR(CDCl₃) [ppm]: 1.35 (t, J=7 Hz, 12H), 3.50 (t, J=22 Hz, 1H),4.05-4.65 (m, 8H), 6.95-7.20 (m, 2H), 7.25-7.70 (m, 3H), 7.90-8.05 (m,1H), 8.95 (brs, 1H)

IR (KBr) [cm⁻¹ ]: 3148, 2984, 1626, 1392, 1232, 1212, 1027 MASS (FAB)m/z: 463 (M+H)⁺ EA (as C₁₉ H₂₈ O₇ P₂ S) Calculated (%): C 49.35 H 6.12Found (%): C 49.39 H 6.11

Example 6 6-Hydroxy-2-naphthylthiomethanediphosphonic Acid (6) ##STR15##

Following the same method as in Example 2, 7.15 g (15 mmol) of thetetraethyl 6-hydroxy-2-naphthylthiomethanediphosphonate obtained inExample 5 was treated with trimethylsilyl bromide, and then hydrolyzedto obtain 4.21 g of the title compound as white crystals.

The yield was 75%.

m.p.: 210°-211° C. ¹ H-NMR (D₂ O) [ppm]: 3.51 (t, J=20 Hz, 1H),7.02-7.13 (m, 2H), 7.48-7.55 (m, 1H), 7.55-7.62 (m, 1H), 7.63-7.69 (m,1H), 7.90-7.97 (m, 1H)

IR (KBr) [cm⁻¹ ]: 3570, 3164, 1636, 1506, 1135, 1046, 939, 919 MASS(FAB) m/z: 351 (M+H)⁺ EA (as C₁₁ H₁₂ O₇ P₂ S) Calculated (%): C 37.73 H3.46 Found (%): C 37.80 H 3.55

Example 7 Tetraethyl 1-naphthylthiomethanediphosphonate (7) ##STR16##

(a) 1,1'-dinaphthyl disulfide

Under an argon atmosphere, to a solution of 22.67 g (100.0 mmol) of1-naphthalenesulfonyl chloride in dry methylene chloride (250 ml) wasslowly added 100.0 g (500.0 mmol) of iodotrimethylsilane, and themixture was stirred for 6 hours. The resulting mixture was poured intoan saturated aqueous solution of sodium bicarbonate, and extracted withmethylene chloride (3×150 ml). The organic layer was washed with asaturated aqueous solution of sodium thiosulfate until the iodinecoloring disappeared, and further washed with water and saturatedsaline, and then dried over anhydrous magnesium sulfate. The solvent wasdistilled off under reduced pressure, and the obtained crystals wererecrystallized from ethyl acetate/n-hexane to obtain 12.43 g of thetitle compound as pale yellow crystals. The yield was 78%.

m.p.: 85°-86° C.

¹ H-NMR (CDCl₃) [ppm]: 7.15-8.00 (m, 12H), 8.20-8.45 (m, 2H)

(b) Tetraethyl 1-naphthylthiomethanediphosphonate

Following the same method as described in Example 1, from 10.09 g (35mmol) of tetraethyl methanediphosphonate and 11.15 g (35 mmol) of1,1'-dinaphthyl disulfide was obtained 9.38 g of the title compound as apale yellow oil. The yield was 60%.

¹ H-NMR (CDCl₃) [ppm]: 1.30 (t, J=7 Hz, 12H), 3.55 (t, J=21 Hz, 1H),3.95-4.45 (m, 8H), 7.30-7.75 (m, 3H), 7.75-8.10 (m, 3H), 8.55-8.75 (m,1H)

IR (KBr) [cm⁻¹ ]: 3434, 2984, 2934, 2910, 1506, 1444, 1.255, 1164, 1098,1013, 971 MASS (FAB) m/z: 447 (M+H)⁺ EA (as C₁₉ H₂₈ O₆ P₂ S) Calculated(%): C 51.12 H 6.33 Found (%): C 51.33 H 6.19

Example 8 1-Naphthylthiomethanediphosphonic Acid (8) ##STR17##

Following the same method as described in Example 2, 6.70 g (15 mmol) oftetraethyl 1-naphthylthiomethanediphosphonate was treated withtrimethylsilyl bromide, and then hydrolyzed to obtain 3.92 g of thetitle compound as white crystals. The yield was 78%.

m.p.: 241°-242° C. (dec)

¹ H-NMR (CD₃ OD) [ppm]: 3.45 (t, J=21 Hz, 1H), 7.30-7.70 (m, 3H),7.75-8.10 (m, 3H), 8.60-8.80 (m, 1H) IR (KBr) [cm⁻¹ ]: 2910, 2892, 1506,1185, 1141, 1006, 932 MASS (FAB) m/z: 335 (M+H)⁺ EA (as C₁₁ H₁₂ O₆ P₂ S)Calculated (%): C 39.53 H 3.63 Found (%): C 39.44 H 3.70

Example 9 Tetraethyl2-[3-methoxy-4-hydroxy-1-naphtbyl]ethenylidene-1,1-diphosphonate##STR18##

Under an argon atmosphere, 55 ml of dry tetrahydrofuran was cooled to 0°C., and a solution of 20.49 g (108 mmol) of titanium tetrachloride indry methylene chloride (15 ml) was slowly added dropwise thereto over aperiod of 15 minutes. To the resulting mixture were added a solution of15.57 g (54 mmol) of tetraethyl methanediphosphonate in drytetrahydrofuran (40 ml) and a solution of 10.92 g (54 mmol) of3-methoxy-4-hydroxy-1-naphthaldehyde in dry tetrahydrofuran (40 ml), andthe mixture was stirred for 10 minutes. After stirring, a solution of21.85 g (216 mmol) of N-methylmorpholine in dry tetrahydrofuran (40 ml)was slowly added dropwise thereto over a period of 30 minutes so as tomaintain the temperature below 5° C. The resulting mixture was stirredfor 30 minutes and then warmed to room temperature and stirred for 5hours. The reaction mixture was poured into ice water and extracted withethyl acetate (3×150 ml). The organic layer was successively washed witha saturated aqueous solution of sodium bicarbonate, with water and withsaturated saline, and then dried over anhydrous magnesium sulfate, afterwhich the solvent was distilled off under reduced pressure. The obtainedresidue was purified by column chromatography (developing solvent -ethanol:ethyl acetate=5:95), and then recrystallized from mixed solventof ethyl acetate/diethyl ether, to obtain 18.37 g of the title compoundas yellow crystals. The yield was 72%.

m.p.: 115.5°-116.5° C.

¹ H-NMR (CD₃ Cl) [ppm]: 1.02 (t, J=7 Hz, 6H), 1.42 (t, J=7 Hz, 6H),3.30-4.50 (m, 8H), 4.04 (s, 3H), 6.90 (brs, 1H), 7.30-7.65 (m, 2H),7.70-7.95 (m, 1H), 8.10-8.35 (m, 1H), 8.20 (s, 1H), 8.35 (dd, J=24.46Hz, 1H)

IR (KBr) [cm⁻¹ ]: 3190, 2990, 1553, 1363, 1247, 1226, 1036, 996 MASS(FAB) m/z: 473 (M+H)⁺ EA (as C₂₁ H₃₀ O₈ P₂) Calculated (%): C 53.39 H6.41 Found (%): C 53.33 H 6.50

Example 102-[3-Methoxy-4-hydroxy-1-naphthyl]ethenylidene-1,1-diphosphonic Acid##STR19##

Following the same method as described in Example 2, 9.45 g (20 mmol) ofthe tetraethyl2-[3-methoxy-4-hydroxy-1-naphthyl]ethenylidene-1,1-diphosphonateobtained in Example 9 was treated with trimethylsilyl bromide, and thenhydrolysis was effected to obtain 4.97 g of the title compound as yellowcrystals. The yield was 69%.

m.p.: 123°-124° C. (dec)

¹ H-NMR (CD₃ OD) [ppm]: 4.01 (s, 3H), 7.37-7.52 (m, 2H), 7.85-7.93 (m,1H), 8.12 (s, 1H), 8.15-8.23 (m, 1H), 8.73 (dd, J=28.46 Hz, 1H)

IR (KBr) [cm⁻¹ ]: 3450, 1603, 1574, 1363, 1125, 1058, 1009 MASS (FAB)m/z: 361 (M+H)⁺ EA (as C₁₃ H₁₄ O₈ P₂) Calculated (%): C 43.35 H 3.93Found (%): C 43.20 H 3.82

Example 11 Tetraethyl2-[3-methoxy-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonate ##STR20##

Under an argon atmosphere, to a solution of 8.50 g (18 mmol) of thetetraethyl2-[3-methoxy-4-hydroxy-1-naphthyl]ethenylidene-1,1-diphosphonateobtained in Example 9 in dry tetrahydrofuran (120 ml) was slowly added2.72 g (72 mmol) of sodium borohydride. The resulting mixture was warmedto 50° C. and stirred for 30 minutes. Next, the mixture was cooled to 0°C., a saturated aqueous solution of ammonium chloride was added theretountil hydrogen was no longer evolved, and the mixture was neutralizedwith 1N hydrochloric acid, and then extracted with ethyl acetate (3×150ml). The organic layer was successively washed with a saturated aqueoussolution of sodium bicarbonate, with water and with saturated saline,and then dried over anhydrous magnesium sulfate, after which the solventwas distilled off under reduced pressure. The obtained residue waspurified by column chromatography (developing solvent-ethanol:ethylacetate=5:95), to obtain 8.28 g of the title compound as a pale yellowoil. The yield was 98%.

¹ H-NMR (CD₃ Cl) [ppm]: 1.22 (t, J=7 Hz, 6H), 1.24 (t, J=7 Hz, 6H), 2.87(tt, J=6.23 Hz, 1H), 3.70-4.50 (m, 8H), 3.71 (dt, J=6.16 Hz, 2H), 3.95(s, 3H), 6.60 (s, 1H), 7.30-7.57 (m, 2H), 7.35 (s, 1H), 7.95-8.05 (m,1H), 8.05-8.15 (m, 1H)

IR (KBr) [cm⁻¹ ]: 3248, 2986, 1630, 1603, 1586, 1479, 1367, 1247, 1025,975 MASS (FAB) m/z: 475 (M+H)⁺ EA (as C₂₁ H₃₂ O₈ P₂) Calculated (%): C53.17 H 6.81 Found (%): C 53.23 H 6.96

Example 12 2-[3-Methoxy-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonicAcid ##STR21##

Following the same method as described in Example 2, 7.12 g (15 mmol) ofthe tetraethyl2-[3-methoxy-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonate obtained inExample 11 was treated with trimethylsilyl bromide, and then hydrolysiswas effected to obtain 3.64 g of the title compound as pale yellowcrystals. The yield was m.p.: 231°-232° C. (dec)

¹ H-NMR (CD₃ OD) [ppm]: 2.68 (tt, J=6.23 Hz, 1H), 3.65 (dt, J=6.16 Hz,2H), 3.96 (s, 3H), 7.33-7.40 (m, 2H), 7.42 (s, 1H), 8.04-8.11 (m, 1H),8.11-8.18 (m, 1H)

IR (KBr) [cm⁻¹ ]: 3324, 2906, 1638, 1475, 1402, 1278, 1176, 1033 MASS(FAB) m/z: 363 (M+H)⁺ EA (as C₁₃ H₁₆ O₈ P₂) Calculated (%): C 43.11 H4.46 Found (%): C 43.02 H 4.45

Example 13 Tetraethyl2-[3-methylthio-4-hydroxy-1-naphthyl]ethenylidene-1,1-diphosphonate##STR22##

Following the same method as described in Example 9, from 14.42 g (50mmol) of tetraethyl methanediphosphonate and 10.91 g (50 mmol) of3-methylthio-4-hydroxy-1-naphthaldehyde was obtained 18.32 g of thetitle compound as yellow crystals. The yield was 75%.

m.p.: 106°-107° C.

¹ H-NMR (CD₃ Cl) [ppm]: 1.03 (t, J=7 Hz, 6H), 1.45 (t, J=7 Hz, 6H), 2.43(s, 3H), 3.70-4.50 (m, 8H), 7.50 (s, 1H), 7.50-7.70 (m, 2H), 7.70-7.90(m, 1H), 8.14 (s, 1H), 8.20-8.45 (m, 1H), 8.80 (dd, J=28.46 Hz, 1H)

IR (KBr) [cm⁻¹ ]: 2990, 1568, 1396, 1313, 1251, 1212, 1046, 982 MASS(FAB ) m/z: 489 (M+H)⁺ EA (as C₂₁ H₃₀ O₇ P₂ S) Calculated (% ): C 51.64H 6.20 Found (% ): C 51.55 H 6.33

Example 14 Tetraethyl2-[3-methylthio-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonate ##STR23##

Following the same method as described in Example 11, 8.79 g (18 mmol)of tetraethyl2-[3-methylthio-4-hydroxy-1-naphthyl]ethenylidene-1,1-diphosphonate wasreduced using 2.72 g (72 mmol) of sodium borohydride to obtain 8.73 g ofthe title compound as a pale yellow oil. The yield was 99%.

¹ H-NMR (CD₃ Cl) [ppm]: 1.22 (t, J=7 Hz, 6H), 1.26 (t, J=7 Hz, 6H), 2.37(s, 3H), 2.82 (tt, J=6.23 Hz, 1H), 3.65 (dt, J=6.16 Hz, 2H), 3.80-4.35(m, 8H), 7.23 (s, 1H), 7.35-7.70 (m, 2H), 7.55 (s, 1H), 7.93-8.18 (m,1H), 8.18-8.42 (m, 1H)

IR (KBr) [cm⁻¹ ]: 3216, 2986, 2928, 1572, 1450, 1388, 1249, 1019, 975MASS (FAB) m/z: 491 (M+H)⁺ EA (as C₂₁ H₃₂ O₇ P₂ S) Calculated (%): C51.42 H 6.60 Found (%): C 51.66 H 6.73

Example 15 2-[3-Methylthio-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonicAcid ##STR24##

Following the same method as described in Example 2, 7.36 g (15 mmol) oftetraethyl 2-[3-methylthio-4-hydroxy-1-naphthyl]ethane-1,1-diphosphonatewas treated with trimethylsilyl bromide, and then hydrolysis waseffected to obtain 4.03 g of the title compound as pale yellow crystals.The yield was 71%.

m.p.: 195°-196° C. (dec)

¹ H-NMR (CD₃ OD) [ppm]: 2.39 (s, 3H), 2.68 (tt, J=6.23 Hz, 1H), 3.63(dt, J=6.16 Hz, 2H), 7.35-7.70 (m, H), 7.57 (s, 1H), 8.00-8.40 (m, 2H)

IR (KBr) [cm⁻¹ ]: 3386, 1578, 1392, 1276, 1210, 1162, 1079, 1013 MASS(FAB) m/z: 379 (M+H)⁺ EA (as C₁₃ H₁₆ O₇ P₂ S) Calculated (%): C 41.28 H4.27 Found (%): C 41.33 H 4.38

Example 16 Adjuvant Arthritis Test

When a tubercule bacillus adjuvant is injected into rats, multiplearthritis, similar to human chronic articularheumatism, is induced. Theanti-inflammatory, anti-rheumatic and bone metabolism-improving effectsof the compounds of the present invention were investigated according tothe following procedure using this adjuvant arthritis model.

0.1 mg of dry, dead tubercule bacilli (Mycobacterium butyricum) cellswas suspended in 0.1 ml of liquid paraffin, and intracutaneouslyinjected into the left bind paw of 7-week-old female Lewis rats. Thecompounds obtained in the Examples were dissolved in sterilizeddistilled water, and subcutaneously administered at a proportion of 20mg per kilogram of weight for 2 consecutive weeks, from the 8th to the21st day after injection of the adjuvant. During that time, the volumesof the left and right paws of the rats were measuredplethysmographically, and edema density was calculated according to thefollowing equation. ##EQU1##

Furthermore, inhibition rate of edema was determined according to thefollowing equation, and are shown in Table 1. ##EQU2##

The rats were sacrified on day 22, and soft X-ray radiographs were takenof the left and right hind legs. The degree of osteoclasia at 5locations on the left and right hind legs was evaluated based on thesoft X-ray radiographs and assigned points from a system of 5 grades,and the total thereof was used as the osteoclasia rating. Furthermore,the inhibition rate of osteoclasia were calculated according to thefollowing equation, and are shown in Table 1. ##EQU3##

The obtained results were indicated with "***" in cases where, based onthe Student t-test and the Tukie multiple comparison method, the levelof significance was P<0.001, with "**" in cases where the level ofsignificance was P<0.01, and with "*" in cases where the level ofsignificance was P<0.05 with respect to the control group to which wasadministered sterilized distilled water alone.

As is clear from Table 1, foot edema and osteoclasia due to primary andsecondary inflammation of adjuvant arthritis were suppressed byadministration of the compounds according to the present invention.

                                      TABLE 1                                     __________________________________________________________________________                                   Inhibition rate                                            Inhibition rate of edema with                                                                    of osteoclasia                                             respect to control group                                                                         with respect to                                       Number                                                                             (%)                control group                                         of   16th or 17th day                                                                       21th day  (%)                                            Compounds                                                                            cases                                                                              Left                                                                              Right                                                                              Left Right                                                                              22nd day                                       __________________________________________________________________________    Compound of                                                                          6    76.8***                                                                           35.2**                                                                             94.5***                                                                            44.3***                                                                            75.2**                                         Example 2                                                                     Compound of                                                                          6    80.0***                                                                           40.8**                                                                             93.0***                                                                            33.4***                                                                            55.2**                                         Example 4                                                                     Compound of                                                                          6    85.6***                                                                           31.8*                                                                              117.3***                                                                           39.9**                                                                             60.3**                                         Example 6                                                                     __________________________________________________________________________

The measurement of foot edema was made on the 17th and 21st days for thecompounds in Examples 2 and 4, and on the 16th and 21st days for thecompound in Example 6.

Example 17 Effect Against Production of IL-1 by Mouse Macrophage CellLine J774-1

Macrophages, one type of lymphocytes, ingest invading microorganisms,blood cell fragments, etc., present antigens to B cells, and releaseactive oxygen to digest foreign bodies, as a foreign body-removalmechanism. At this time the macrophages release a number of cytokines,including IL-1 which causes fever, inflammation, chondroclasia,osteoclasia, activation of leukocytes, damage to vascular endothelialcells, etc., and is also known to exhibit various effects by inducingthe production of other cytokines.

Mouse macrophage cell line J774-1 is selected from cells exhibiting highproduction of IL-1, and it is known to produce IL-1 when stimulated byLPS. With this cell line, the inhibitory effects to IL-1 production ofthe compounds according to the present invention were determined by thefollowing procedure.

J774-1 cells were cultured in an RPMI-1640 culture medium containing 10%fetal calf serum and 50 μM of 2-mercaptoethanol, and prepared to a cellconcentration of 2×10⁵ cells/ml. The cell suspension was distributedinto a 24-well plate to 1 ml per well, and cultured for 30 minutes. LPSwas then added thereto to a final concentration of 1 μg/ml, and at thesame time the compounds obtained in the Examples dissolved in sterilizeddistilled water were added to a concentration of 100 μM. After 24 hours'culturing at 37° C. in a 5% CO₂ environment, the supernatant wasrecovered, centrifuged to remove the cell fragments, etc., and thenpassed through a 0.22 μm filter for sterilization.

The measurement of IL-1 activity was made by measuring the proliferationof thymocytes in male C3H/He J mice. In fact, 4 to 6-week-old maleC3H/He J mouse was used, and the thymus was taken. The thymus wasdissociated in an RPMI-1640 culture medium containing 10% fetal calfserum and 50 μM of 2-mercaptoethanol, and a cell suspension was preparedto a concentration of 2×10⁷ cells/ml. Phytohemagglutinin was added tothe cell suspension to a final concentration of 1%, and this was used asthe T cell suspension.

A two-fold serial dilution was made of the above obtained sample in a96-well multiplate to a volume of 50 μl per well, and 50 μl of the Tcell suspension was added to each well. The T cells were cultured for 72hours, and the IL-1 activity was determined by the rate of cellproliferation. The cell proliferation was calculated using as the valueof the absorbance at 570 nm of the pigment produced upon reduction of3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide by themitochondria of the viable cells, added at 4 hours prior to completionof the culturing, with 100% proliferation defined as the maximumproliferation of T cells induced by recombinant human IL-1, 0% definedas proliferation with no addition of IL-1, and the units of the sampledefined as the degree of dilution of the sample which caused 50%proliferation of the T cells.

At this time, the inhibition rates of the compounds according to thepresent invention against IL-1 production by J774-1 cells stimulatedwith 1 μg/ml of LPS was calculated using the following equation. Theresults are shown in Table 2. ##EQU4##

                  TABLE 2                                                         ______________________________________                                                      IL-1 production inhibition rate                                               with respect to control group                                   ______________________________________                                        Compound of Example 2                                                                         41.7%                                                         ______________________________________                                    

Example 18

Rabbit cartilage cells are separated from knee joint and cultured. Whenthey are stimulated with IL-1, proteoglycan, the main constituentglycoprotein of cartilage cells, is degraded. Using this consequence asan index, the IL-1-inhibiting effects of the compounds according to thepresent invention were determined in the following manner.

Three-week-old male New Zealand white rabbits weighing 250 g-300 g weresacrified under diethyl ether anesthesia, and their knee joints wereseparated. The cartilaginous sections were cut out from the knee jointwith a scalpel, and then immersed in CMF solution containing 0.14Msodium chloride, 4 mM potassium chloride, 0.4 mM sodium dihydrogenphosphate, 12 mM sodium bicarbonate, 0.2 mM potassium dihydrogenphosphate and 11 mM glucose. The cartilage was placed in 0.1% EDTA andincubated at 37° C. for 20 minutes. The supernatant was removed off,0.15% trypsin was added to the mixture, and then that was incubated at37° C. for 60 minutes. The mixture was washed 3 times with CMF solution,and then placed in 15% collagenase and further treated at 37° C. for 105minutes. The cartilage cells were isolated from the cartilaginous tissuefragments by pipetting and pushed through 120 μm nylon mesh, and thensubjected to centrifugation at 4° C., 500 g for 7 minutes to obtain thecartilage cells. The cells were washed 3 times, and then suspended inDulbecco MEM culture medium containing 10% fetal calf serum, toconcentration of 1.2×10⁵ cells/ml. The cells were distributed into a48-well plate to 250 μl per well and cultured for 5 days untilconfluence. Then, the culture solution was exchanged with Dulbecco MEMmedium containing 0.3% fetal calf serum. After the further incubationfor 24 hours, S-labelled inorganic sulfuric acid was added toconcentration of 185 kilobecquerels, and the incubation was continuedfor further 24 hours. The cells were washed 3 times with Dulbecco MEMmedium, the culture medium was exchanged with a BGjb medium containing0.1% bovine serum albumin, and recombinant human IL-1β was added to aconcentration of 30 units/mi. At the same time, the compounds accordingto the present invention were dissolved in sterilized distilled water,and added to a final concentration of 100 μM. At 24 hours after IL-1stimulation, the culture supernatant and the cell layer were collected.

The cell layer was decomposed by adding 200 μg of Pronase E andtreatment at 37° C. for 24 hours. To the culture supernatant weresuccessively added 0.05 ml of 0.1 mg/ml chondroitin sulfate, 0.5 ml of 2mM magnesium sulfate, 0.5 ml of a buffer solution (pH 7.8) containing 5mM calcium chloride and 0.2M Tris-HCl and 0.5 ml of a solution of 1%cetyl pyridinium chloride and 20 mM sodium chloride, and theproteoglycan which precipitated upon treatment at 37° C. for 2 hours wascollected into a glass filter, a liquid scintillator was added thereto,and the count was made using a liquid scintillation counter.

To the digested cell layer solution were successively added 0.05 ml of0.1 mg/ml chondroitin sulfate, 0.5 ml of 2 mM magnesium sulfate and 0.5ml of a solution of 1% cetyl pyridinium chloride and 20 mM sodiumchloride, and the proteoglycan which precipitated upon treatment at 37°C. for 2 hours was harvested into a glass filter, a liquid scintillatorwas added thereto, and the count was made using a liquid scintillationcounter.

Each of the obtained counts was divided by the count at the initialaddition of inorganic sulfuric acid, and expressed as a percentage. Theobtained results were indicated with "$$" in cases where, using theStudent ttest, the level of significance was P<0.01 with respect to thenon-stimulated control group and with "**" in cases where the level ofsignificance was P<0.01 with respect to the IL-1-stimulated controlgroup. As shown in Table 3, the compounds according to the presentinvention inhibited the degradation of proteoglycan from theIL-1-stimulated cell layer, and are thus effective as IL-1-inhibitingagents.

                  TABLE 3                                                         ______________________________________                                                   Supernatant Cell layer                                             ______________________________________                                        Not stimulated                                                                             0.63 ± 0.035                                                                             0.45 ± 0.036                                    IL-1-stimulated                                                                             1.06 ± 0.018$$                                                                           0.11 ± 0.004$$                                 IL-1-stimulated and                                                                        0.78 ± 0.015**                                                                           0.31 ± 0.011**                                  compound-treated                                                              (Compound of                                                                  Example 2)                                                                    ______________________________________                                    

Example 19

Neutrophils are known to ingest foreign bodies for their removal and toproduce active oxygen and digestive enzymes, as a biological defensemechanism. However, during chronic inflammation, etc., the active oxygenand digestive enzymes produced by neutrophils also damage normal tissue,and are thought to further reinforce inflammation. Here, the effects ofthe compounds according to the present invention against the release ofactive oxygen from neutrophils were determined in the following manner.

Using 3.8% sodium citrate as an anticoagulant, 50 ml of blood was takenfrom a human vein. The blood was mixed with the same volume of asolution of 2% dextran and physiological saline, and the mixture wasshaken several times and then allowed to stand at 37° C. for 30 minutes.The upper layer was separated off and overlayered onto the same volumeof a Ficoll-Paque solution. The precipitate resulting from 30-minutes'centrifugation at 20° C. 1400 rpm was taken, the cells were resuspendedin Hanks' balanced salt solution, centrifugation was performed at 20°C., 1000 rpm for 5 minutes, and the precipitated cells were washed. Thecontaminating erythrocytes were eliminated by subjection to osmoticshock, and finally the neutrophils were suspended in Hanks' balancedsalt solution to a concentration of 1×10⁶ cells/ml. Of theseneutrophils, 1×10⁵ cells were incubated at 37° C. with 10⁹ M of thestimulant, formyl-methionyl-leucyl-phenylalanine (fMLP), and at the sametime the compounds according to the present invention were added theretoand the production of active oxygen was measured. For the measurement ofthe active oxygen, 2-methyl-6-phenyl-3,7-dihydroimidazo[1,2a]pyrazine-3-one (CLA) was reacted therewith resulting in excitedcarbonyl compounds, and utilizing the phenomenon whereby light isemitted at 380 nm during their transition to ground state, the maximumluminescence intensity was measured with a luminometer. The inhibitionrate against active oxygen production was calculated according to thefollowing equation. The results are shown in Table 4. ##EQU5##

                  TABLE 4                                                         ______________________________________                                                          Active oxygen                                                                 production inhibition rate                                  ______________________________________                                        Compound of Example 2                                                                       10 μm  29.4 ± 6.58%                                       ______________________________________                                    

Example 20

In conditions of osteoporosis, it is thought that the balance betweenbone formation and bone resorption is lost, with bone resorption beingaccelerated. Bone resorption is thought to occur due to the activationand increase in the number of osteoclasts, and a model thereof is anexperiment in which mouse osteocytes are planted on dentin slices,causing bone resorption due to the stimulation of active-type vitaminD₃. Using this model, the bone resorption-inhibiting effects of thecompounds according to the present invention were determined.

The femurs and tibias were separated from 10 to 15- day-old ICR mice,and minced in an α-MEM culture medium containing 5% fetal calf serum,and an osteoclast suspension was prepared containing bone marrow cellsand bone matrix. The large bone fragments were removed using a nylonmesh, and the viable cells were stained using trypan blue staining,while the osteoclasts were stained using tartaric acid-resistant acidicphosphatase staining, and a cell suspension was prepared which containedthe osteoclasts at a proportion of about 0.05-0.1%. The dentin was cutto thicknesses of 150 μm using a low-speed rotating diamond cutter, andpunched with a puncher to the size of wells of a 96-well plate. Thedentin slices were placed in a 96-well plate, and the cell suspensionprepared as described above was placed thereupon to a concentration of500 osteoclasts per well. As a stimulant, 10 nM active-type vitamin D₃was added thereto, and at the same time the preparations according tothe present invention were added thereto to concentrations of 10 μM and100 μM. The cells were cultured at 37° C. in 10% CO₂ environment, andafter 4 days' culture the resorption pits which formed on the dentinslices were stained with hematoxylin, and then observed under amicroscope and counted. The rate of inhibition of resorption pitformation was calculated according to the following equation. ##EQU6##

The results are shown in Table 5. The results were indicated with "*" incases where, based on statistical calculation using the Student t-test,the level of significance was P<0.05 and with "**" in cases where thelevel of significance was P<0.01 with respect to the active-type vitaminD₃ -stimulated control group.

                  TABLE 5                                                         ______________________________________                                                           Inhibition rate with                                                          respect to control                                                            (%)                                                        ______________________________________                                        Compound of Example 2                                                                          10 μM                                                                              18.8 ± 6.23                                                       100 μM                                                                              92.1 ± 0.43**                                     Compound of Example 4                                                                          10 μM                                                                              47.7 ± 2.18*                                                      100 μM                                                                              94.8 ± 1.04**                                     Compound of Example 6                                                                          10 μM                                                                              30.6 ± 12.5                                                       100 μM                                                                              90.8 ± 1.33**                                     ______________________________________                                    

[Industrial Applicability]

The compounds according to the present invention possess anti-IL-1,anti-oxidation and anti-bone resorption effects, etc. and are thususeful as anti-inflammatory agents, analgesics, antirheumatic agents,agents for bone metabolism disorders, agents for autoimmune diseases,agents for infections, agents for dermatologic diseases, antiallergicagents, antioxidants and therapies for ischemic organ damage.

We claim:
 1. A methanediphosphonic acid derivative represented by the general formula (I): ##STR25## wherein X and Y represent substitution groups on the naphthyl group, and represent a halogen atom, nitro group, nitrile group, alkyl group, alkoxy group, trifluoromethyl group, the group: ##STR26## provided that Z¹ and Z² represent, independently of each other, a hydrogen atom or an alkyl group, or Z¹ or Z² may form a ring comprising carbon atoms or a ring comprising carbon atoms and hetero atoms, the group: ##STR27## provided that Z¹ and Z² are the same as above, and Z³ represents oxygen or sulfur, thiol group, hydroxyl group, alkylthio group, arylthio group, acyloxy group, acylamino group, acylthio group, acyl group, alkenyl group, aryl group, cycloalkyl group, COOH group or COO-alkyl group; m represents an integer of 0 to 3; n represents an integer of 0 to 4; and each X of the (X)_(m) and each Y of the (Y)n may be either identical or different; . . . represents a double bond or single bond; A is --(CH₂)a--(D)b--(CH₂)c--, wherein D is sulfur, NH, alkyl-substituted N or CH₂, a and c are integers of 0 to 10 and b is 0 or 1, or --(CH═CH)d--CH═, wherein d is an integer of 0 to 2, and B does not exist when A represents --(CH═CH)d--CH═, B refers to a hydrogen atom, alkyl group, amino group, monoalkylamino group, dialkylamino group, acylamino group, alkoxy group, trialkylsiloxy group or acyloxy group, and each of R¹, R², R³ and R⁴ is the same or different and is a hydrogen atom, straight or branched alkyl group having 1 to 7 carbon atoms, or a pharmacologically allowed cation; with the proviso that when D is sulfur, X is not a hydroxyl group and both a and c are 0; and with the proviso that when D is --CH₂ --, or when d is 0 or 1, X is not an alkyl group and m is 2 to
 3. 2. The methanediphosphonic acid derivative set forth in claim 1 wherein the naphthyl group is a 1-naphthyl or 2-naphthyl group.
 3. A methanediphosphonic acid derivative which is selected from the group consisting of tetraethyl 2 -naphthylthiomethanediphosphonate, 2 -naphthylthiomethane, diphosphonic acid, tetraethyl 6 -methoxy-2-naphthylthiomethanediphosphonate, 6-methoxy-2-naphthylthiomethanediphosphonic, acid, tetraethyl 6-hydroxy-2-naphthylthiomethanediphosphonate, 6-hydroxy-2-naphthylthiomethanediphosphonic, acid, tetraethyl 1-naphthylthiomethanediphosphonate, 1-naphthylthiomethanediphosphonic acid, tetraethyl 2-(3-methoxy-4-hydroxy-1-naphthyl)ethenylidene-1,1-diphosphonate, 2-(3-methoxy-4-hydroxy-1-naphthyl)ethenylidene-1,1-diphosphonic acid, tetraethyl 2-(3-methoxy-4-hydroxy-1-naphthyl)ethane-1,1-diphosphonate, 2-(3-methoxy-4-hydroxy-1-naphthyl)ethane-1,1-diphosphonic acid, tetraethyl 2-(3-methylthio-4-hydroxy-1-naphthyl) ethenylidene-1,1-diphosphonate, tetraethyl2-(3-methylthio-4-hydroxy-1-naphthyl)ethane-1,1-diphosphonate, and 2-(3-methylthio-4-hydroxy-1-naphthyl)ethane-1,1-diphosphonic acid.
 4. The methanediphosphonic acid derivative set forth in claim 1, which is 2-naphthylthiomethane diphosphonic acid.
 5. The methanediphosphonic acid derivative set forth in claim 3, which is 6 -methoxy-2 -naphthylthiomethanediphosphonic acid.
 6. The methanediphosphonic acid derivative set forth in claim 3, which is 6-hydroxy-2-naphthylthiomethanediphosphonic acid.
 7. A pharmaceutical composition comprising a methane diphosphonic acid derivative set forth in claim 1 or 2, and a pharmaceutical acceptable carrier.
 8. A method for the inhibition of inflammation comprising administering to a subject a methane diphosphonic acid derivative set forth in claim 1 or
 2. 9. A method for the treatment of rheumatism comprising administering to a subject a methane disphosphonic acid derivative set forth in claim 1 or
 2. 10. A method for the treatment of bone metabolic diseases comprising administering to a subject a methane diphosphonic acid derivative set forth in claim 1 or
 2. 11. A method for the inhibition of interleukin-1 production or interleukin-1 induced cellular response comprising administering to a subject a methane diphosphonic acid derivative set forth in claim 1 or
 2. 12. A method for the inhibition of active oxygen production in neutrophels comprising administering to a subject a methane diphosphonic acid derivative set forth in claim 1 or
 2. 13. A method for the inhibition of bone resorption comprising administering to a subject a methane diphosphonic acid derivative set forth in claim 1 or
 2. 